Abstract
In this study, we characterized the small subunit of ubiquinol-cytochrome C reductase (Bmuccr) of the silkwormBombyx mori, a model insect of Lepidopteron species. The Bmuccr gene covers a 1.4 kb genome region and contains 3exons. The ORF contained 354bp and encoded 117 amino acid residues, which shares 69% overall amino acid sequenceidentities with the subunit VII of ubiquinol-cytochrome C reductase from bovine. Phylogenetic tree showed Bmuccr hadhigh homology with T. castaneum homologous. The multiple sequence alignment of 16 subunit VII homologues showsthat Bmuccr is very hydrophilic, has a characteristic charge distribution, and has a high helical content. Expressionanalysis indicated that Bmuccr was highly expressed in larva stage and was down-regulated in embryos stage and adultstage of silkworm. The tissue-specific expression indicated Bmuccr had high-expression level in tissues that consumeoxygen. The analysis of domain structure of this protein suggested that it might be involved in correct assembly of thecytochrome bcl complex. Definition of the homologous of bovine subunit VII of ubiquinol-cytochrome C reductaseshould facilitate further analysis of structure/function relationships of silkworm cytochrome bcl complex.
Highlights
Polypeptide subunits of bovine ubiquinol-cytochrome C reductase complex, 7.2kDa protein maintains contact with cytochrome c1 and iron sulfur protein (Schagger et al, 1983, p. 307)
The Bombyx mori ubiquinol-cytochrome C reductase 14kDa subunit (Bmuccr) cDNAs without any mismatch nucleotides were cloned into the expression vector PET30a (Novagen) and were transformed into the E. coli expression strain BL21 (DE3)
The crystal structure of subunit VII of bovine cytochrome bcl complex is available in PDB (PDB ID: 1bccF) and it was used as the template to build the model of Bmuccr protein (Figure 2B)
Summary
The cytochrome bcl complex (commonly known as ubiquinol-cytochrome c reductase or Complex III) is a segment of the mitochondrial respiratory chain that catalyzes reduction of Cyt c by the oxidation of ubiquinol (Hatefi et al, 1967, p. 235; Bechmann et al, 1992, pp.199). Two large "core" proteins, three respiratory proteins that directly participate in electron transport (Cyt b, Cyt cl, and the "Rieske" iron sulfur protein), and four to six small proteins with molecular masses of less than 20 kDa. Since bacterial bc1complexes that contain only the respiratory subunits have the same activity as the eukaryotic enzymes (Trumpower, 1990, p.101), the role of the small subunits is not quite understood. Polypeptide subunits of bovine ubiquinol-cytochrome C reductase complex, 7.2kDa protein (subunit X) maintains contact with cytochrome c1 and iron sulfur protein We report the isolation, tissue expression, and in vitro expression of a novel silkworm cDNA which encodes 117 amino acids that share 69% sequence identity with subunit VII of the bovine bc complex
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